Summary: Real-time PCR, also known as kinetic PCR, qPCR, qRT-PCR and RT-qPCR, is quantitative PCR method for the determination of cope number of PCR templates such as DNA or cDNA in a PCR reaction. There are two flavors of real-time PCR: probe-based and intercalator-based. Both methods require a special thermocycler equipped with a sensitive camera that monitors the fluorescence in each well of the 96-well plate at frequent intervals during the PCR Reaction. Probe-based real-time PCR, also known as TaqMan PCR, requires a pair of PCR primers as regular PCR does, an additional fluorogenic probe which is an oligonucleotide with both a reporter fluorescent dye and a quencher dye attached. Intercalator-based method, also known as SYBR Green method, requires a double-stranded DNA dye in the PCR reaction which binds to newly synthesized double-stranded DNA and gives fluorescence. TaqMan method is more accurate and reliable than SYBR green method, but also more expensive.

SYBR Green Quantitative PCR Protocol(Lusis Lab, UCLA)Quantitative PCR is a method used to detect relative or absolute gene expression level. All qPCR involves the use of fluorescence to detect the threshold cycle (Ct) during PCR when the level of fluorescence gives signal over the background and is in the linear portion of the amplified curve. This Ct value is responsible for the accurate quantization of qPCR. http://www.genetics.ucla.edu/labs/lusis/greenquant...Added: Sat Mar 21 2009, Hits: 28511, Reviews: 0Write reviewCached

Protocol for Real-Time RT-PCR(Dr. Xiaowei Wang at the Department of Molecular Biology, Massachusetts General Hospital)This protocol describes the detailed experimental procedure for real-time RT-PCR using SYBR Green I. The procedure begins with reverse transcription of total RNA. The cDNA is then used as template for real-time PCR with gene specific primers. You may need to modify this protocol if you use different reagents or instruments for real-time PCR. http://pga.mgh.harvard.edu/primerbank/PCR_protocol...Added: Fri May 14 2004, Hits: 2091, Reviews: 0Write reviewCached

Quantification using real-time PCR technology
(Dr. Michael W. Pfaffl, Gene Quantification )The introduction of real-time PCR technology has significantly improved and
simplified the quantification of nucleic acids, and this technology has become an invaluable tool for many scientists working in different disciplines. Especially in the field of molecular diagnostics, real-time PCR-based assays have gained favour in the recent past. However, the wide use of real-time PCR methods has also highlighted some of the critical points and limitations of these assays. http://www.wzw.tum.de/gene-quantification/klein-20...Added: Sun Sep 22 2002, Hits: 13475, Reviews: 0Write review